技术资料

Neural crest cells contribute to craniofacial formation by differentiating into skeletogenic mesenchyme and neuro-glial lineages. Using Smart-seq2 single-cell transcriptomics,we show that mesenchymal fate commitment correlates specifically with the expression of rRNA-modifying and ribosome assembly factors,rather than structural ribosomal proteins. Notably,EMG1 and NHP2 introduce key post-transcriptional modifications into 18S rRNA,including m¹acp³ψ at U1248,which requires TSR3 for final maturation. Disrupting NHP2 or TSR3 in vitro and in vivo perturbs cranial neural crest differentiation; post-migratory temporal knockout of Polr1a or Polr1c also causes craniofacial malformations. These findings align with cell type-specific m¹acp³ψ levels during neural crest differentiation. Given the neural crest contribution to neuroblastoma,we analyze patient data to find that elevated ribosomal control and rRNA-modifying proteins predict poorer outcomes. Complementary experiments in neuroblastoma cell lines reveal functional roles for TSR3 and WDR74 in mesenchymal-like tumor states. Together,our results link rRNA modifications and ribosome assembly to fate decisions,suggesting ribosomal heterogeneity shapes both normal development and tumor progression. Neural crest cells differentiate into skeletogenic mesenchyme and neuro-glial lineages,thereby contributing to craniofacial formation. Here,single-cell analysis of cranial neural crest shows that specific rRNA modification and ribosome assembly factors contribute to skeletogenic fate. Their disruption causes craniofacial defects,while high levels in neuroblastoma predict poor survival. View Publication

Understanding how chemical stress perturbs human lung physiology requires models that capture dynamic molecular responses in real time. Here,we established a CRISPR/Cas9-engineered human induced pluripotent stem cell (hiPSC)-derived lung organoid expressing endogenous G3BP1–mCherry,enabling live,non-destructive visualization of stress granule (SG) formation under toxicant exposure. The organoids recapitulated airway and alveolar epithelial diversity and displayed lamellar body-like ultrastructures,indicating advanced maturation. Time-lapse imaging revealed rapid and reversible SG dynamics across chemically distinct stressors,while cytotoxicity assays showed that these organoids are significantly more sensitive than conventional 2D or cancer-derived lung models. Importantly,SG dynamics were linked to exposure duration–dependent changes in epithelial barrier integrity,indicating that SG formation precedes overt epithelial injury and serves as an early indicator of toxicant-induced cellular stress. Integration with high-content screening enabled quantitative,image-based analysis of cellular stress phenotypes,greatly enhancing throughput and mechanistic insight,thereby provided next-generation New Approach Methodologies for lung toxicity assessment. Together,this hiPSC-derived lung organoid SG reporter platform links early molecular stress adaptation to tissue-level responses,offering a predictive and mechanistically informative framework for human-relevant lung toxicity evaluation. View Publication

Background: Lenalidomide is an immunomodulatory drug approved in the treatment of autoimmune disease,inflammation,and cancer. Its impact continues to grow due to its diverse spectrum of effects hampered only by toxicities and reduced efficacy. Therefore,development of strategies that enhance function while reducing drawbacks remains a prime goal. Objective and Hypothesis: The mechanisms of action of lenalidomide on the activity of natural killer cells (NK cells) remains understudied yet could be critical for the development of strategies to enhance its efficacy. These cells are critical drivers of anti-tumor immune responses which are often functionally suppressed in malignancies. NK cell and T cell survival and function is driven by the IL-2 family of cytokines (IL-2 or IL-15) and work has shown that lenalidomide potentially works by increasing the secretion of IL-2 by other lymphocytes,such as CD4+ T helper cells. Thus,we hypothesized that improving NK activity with IL-2 family of cytokines could lead to enhanced lenalidomide-induced responses of these cells. Results: We show that lenalidomide does not affect NK cell viability but reduces their proliferation through cell cycle arrest which could be overcome by exogenous addition of IL-2 family of cytokines. Moreover,lenalidomide induced the secretion of IL-2 on isolated NK cells although it also modulated NK receptor expression,such as NKp46,trough downregulation of PI3K/AKT pathway reduction. This was overcome by exogeneous addition of IL-2 family of cytokines increasing natural cytotoxicity,through higher perforin and granzyme expression. Mechanistically,this increased gene and protein expression occurred through the activation of STAT5 by lenalidomide which was also enhanced through the exogenous addition of IL-2 family of cytokines and modulation of IL-2R subunit changes. Conclusions: These data provide a rationale for the combination of lenalidomide with IL-2 family of cytokines to enhance the effectiveness of NK cells. View Publication